NCBI BLAST:
Identifying Sequences

NCBI BLAST allows you to input a sequence from DNA, RNA or protein residues (amino acids) and find sequences that are identical or similar.

To get to BLAST from the NCBI home page, click BLAST from the Popular Resources menu bar on the right of the page.

[Click on image above to expand]

You can also get to BLAST directly by going to the BLAST home page.

For this simple exercise, we will give you a nucleotide sequence to identify.

Click Nucleotide BLAST on the left of the page.

Click the Next button below.

Identifying Sequences with BLAST

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There are many options on the Standard Nucleotide BLAST (also called "blastn") page.

For example, you can:

• Select different databases to search
• Exclude certain data sources
• Select a specific algorithm by which to search

For your first BLAST, we will keep this very basic. We will use the default options to enter a sequence string. We'll use BLAST to identify the organism it came from and see what else we can learn about it.

Identifying Sequences with BLAST

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Copy and paste the entire string of nucleotide symbols, below, into the box under Enter Query Sequence.

Uncheck the box labeled "Align two or more sequences" if it is checked.

Then scroll down and click the BLAST button.

Identifying Sequences with BLAST

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You may need to be patient.

BLAST is crunching a huge amount of data.

You will see a screen like this for a while during processing:

Reading your BLAST Results

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Once your results are displayed, scroll down to see a header followed by the results of your search. The results can be displayed in several different views, including a list of sequence "Descriptions," via a "Graphic Summary," and via a more detailed "Alignments" view.

Select the Graphic Summary by clicking on this tab:

This shows a graphic summary of the top 100 results.

Reading your BLAST Results

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Each bar in this graph represents a match with another sequence in the database. The color of each line represents the extent to which the sequence in the database aligns with the sequence you input (the "Query" sequence). The higher the alignment score, the closer the alignment.

See the color key:

What are these highly aligned sequences? Where did they come from?

One way to find out is to click on one of the bars in the graphic summary. Try that now.

Exploring your BLAST Results

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You should be viewing your BLAST results in your other browser window.

Click on the Descriptions tab to learn more about each of the sequences that aligned with yours.

Click on the description of the sequence to see the alignment.

For this exercise, select one of the sequences labeled, "Homo sapiens...mitochondrion."

[Note: Your results may look slightly different.]

Clicking on a sequence will bring you to the Alignments view.

Exploring your BLAST Results

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You can now see all the nucleotide base matches between your sequence (the "query" sequence) and the sequence from the database (the "subject" sequence).

This particular alignment isn't very interesting to look at because the two sequences match perfectly. In the next example, we'll look at two sequences that do not perfectly align so that you can look at differences.

Our goal right now is simply to identify the sequence and explore the results.

Exploring your BLAST Results

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To go to the subject sequence in the Nucleotide database, there are several links from the alignment.

The first two, (1) one in the header next to Download labeled GenBank and (2) another link from the Sequence ID, take you to the record for the full sequence as it was submitted (or created). Remember that our match starts around base 7585.

The third link (3), adjacent to the range (also labeled GenBank), takes you to a record displaying just the range of interest (around 7585 to around 8268).

Either record might be useful, but let's look at the record for the entire sequence that was submitted and look at our query sequence in that context.

Follow the link to the GenBank record in the Nucleotide database from your Sequence ID (MZ387763.1 in this example):

[If you wish, you can use this link instead to go to MZ387763.1]

Exploring your BLAST Results

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You should now be in the NCBI Nucleotide database, looking at a record labeled something like, "Homo sapiens haplogroup B2d mitochondrion, complete genome."

[Note: You may be looking at a different record.]

Many of the records we look at in this course are Reference Sequences or "RefSeq" records, which are curated by NCBI. But this is an "original" sequence record submitted by a GenBank participant.

Exploring your BLAST Results

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An interesting part of a Nucleotide record is the section labeled FEATURES. Called the "feature table," this is the part that reflects scientists' annotations -- notes on what biological features of interest are known about a sequence.

Scroll down the feature table of this mitochondrial DNA record. Definitions of some of the feature labels can be found in the Sample GenBank Record.

Two features of major interest include:

• CDS: A coding sequence, or region of nucleotides that corresponds with amino acids in a protein.
• gene: A region identified as a gene. A gene may include multiple sections of coding sequences, so the same nucleotide sequence (shown in a number range) may be labeled as CDS and gene.

In the feature table, each labeled feature is hyperlinked to the sequence itself, which is at the bottom of the record. Click on the first instance of a gene label in this feature table.

You can now see the sequence for the gene highlighted in the context of the rest of the sequence:

Exploring your BLAST Results

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The tools that appear at the bottom provide a useful way to learn and navigate your way around the features.

For example, since you clicked on a gene, you can now toggle through all the genes in this record using the tool in the lower left.

Click around this feature table for a few minutes to get more accustomed to looking at this data.

Conclusion

Congratulations! You've completed this exercise in identifying sequences using NCBI BLAST.

You can now close the NLM Navigator windows.